Alzheimer's disease (AD) is defined in part by the appearance of intracellular inclusions composed of the microtubule associated protein tau. The mechanisms that drive tau aggregation in the highly prevalent sporadic form of AD are not fully understood, but appear to involve abnormal post-translational modifications. To gain insight into the modifications that accompany tau lesion formation in AD, we conducted a preliminary structural analysis of tau aggregates isolated from authentic disease tissue specimens using mass spectrometry methods. Our results revealed that a previously unrecognized tau modification, lysine methylation, copurified with tau aggregates. The preliminary methylation signature involved sites that are known to mediate tau ubiquitylation and other post-translational modifications, suggesting that methylation is a normal tau modification and a candidate for influencing tau lesion formation in disease. We now propose an exploratory project to validate methylation as a pathophysiological tau modification, to place it into the context of modification signatures associated with both normal and aggregated tau isolated from post-mortem human tissue, and to gain preliminary insight into its potential effects on tau aggregation propensity an microtubule polymerization activity. Upon achieving the milestones proposed in this study, the AD field will gain the key information necessary to investigate potential cross-talk among tau post-translational modifications, to guide future identification of methyltransferases and demethylases that act on tau protein, and to identify novel disease-associated epitopes that may aid in the pre-mortem assessment of AD.